ABSTRACT Allergic diseases are caused by inappropriate immune responses to otherwise harmless antigens, and, in one form or another, affect up to 25% of the population in the United States, with a disproportionate increase especially among children with asthma and food-related hypersensitivities. While many immune cell types are implicated in the pathogenesis of allergic disease there is growing evidence that basophil granulocytes play a far greater role than initially thought. Indeed, a good portion of this excitement stems from work done ~20 years ago (including our own) showing that human basophils are a prolific source of IL-4 & IL-13 ?cytokines central to the allergic diathesis. More recently, these findings have been confirmed and extended in vivo using mouse models. Moreover, some mouse models now show evidence of an ?axis? whereby epithelial cell (EC)-derived cytokines (e.g. TSLP, but also IL-33 and IL-25) activate basophils to produce IL-4 (& IL-13) that is seemingly critical for initiating IgE synthesis and thus sensitization. In contrast, confirmatory results identifying TSLP as an activator of human basophils have not been forthcoming. In exploring this translational discrepancy, we have now uncovered a remarkably robust, and potentially unique, mode of EC- dependent activation of human basophils that occurs independently of known EC-derived cytokines. Basophils co-cultured with EC of lung origin (A549 cells) in the presence of IL-3 secrete high IL-4/IL- 13 levels yet to be achieved with other modes of stimulation. This response is preceded by mediator (histamine) release, but with delayed kinetics compared to that seen with standard Fc?RI-dependent activation. New evidence points to an IgE-binding protein on EC (e.g. galectin-3) that is mediating this unique mode of basophil activation. Two aims are proposed to further characterize this novel response. Aim 1 explores the nature of this interaction, hypothesizing the need for cell-to-cell interactions. The role of IL-3 priming of basophils and the requirement for IgE are also to be investigated. We proposed experiments also testing whether EC activate other IgE-bearing cells (e.g. mast cells) for similar activity. And, other EC lines (including normal EC) are to be investigated for mediating basophil activation, hypothesizing that ?danger signals? (viruses, cytokines, TLR agonists) play an important role in enabling the latter to possess a similar capacity. Aim 2 studies are to explore the role of known IgE-binding proteins such as galectin-3 (but also CD23b, and potentially novel ones), in meditating this activity. Overall, our unique experience working with human basophils makes this R21 application highly innovative, and increases the probability that novel, significant, and clinically relevant findings will be discovered.